Lineage and induction of ventrolateral mesoderm

描述（由申请人提供）：在脊椎动物中，许多组织和器官全部或部分来源于中胚层，包括骨骼肌、心脏、骨骼和肾脏。一个复杂的基因网络调节着这些不同的中胚层衍生物的逐渐分化。在非洲爪蟾中，中胚层是由早期原胚赤道周围的一束细胞形成的。在非洲爪蟾的中胚层模式模型中，精子组织者占有重要地位，并被认为通过其诱导因子的分泌来指定中胚层的命运。我们最近发现，在中原肠期，一条与精子组织无关的途径沿着中胚层的动物-植物轴建立了一个模式。在这个轴上可以观察到两个不同的域。动物结构域表达基因brachyury，似乎注定要成为脊索和some - ite，而植物结构域注定要成为腹外侧中胚层。这一建议的目的是进一步阐明在动物/植物轴上建立模式的机制。特别是，我们将研究结生长因子在腹侧中胚层诱导中的作用。我们的初步结果表明，Xenopus nodal-related-2 （Xnr2）表达细胞仅限于原胚胚胎的推定腹侧中胚层。这里提出的谱系追踪实验旨在严格测试这些细胞的命运。最终确定早期假定的中胚层细胞的命运，并将它们与限制性基因表达模式联系起来，是阐明命运规范的机制和时间的重要第一步。我们还假设淋巴结在腹侧中胚层的诱导中起着重要作用，我们将通过使用显性阴性结构和抑制结合蛋白来检查原肠胚中淋巴结的活性。最后，我们的初步结果表明，Xnr2信号和低FGF活性（如存在于植物边缘区）的结合，使外胚层外植体分化为血液（一种腹侧中胚层衍生物），以响应骨形态发生蛋白（BMP）。我们将使用减法和表达克隆方法来鉴定Xnr2信号下游的基因，这些基因负责改变外胚层外植体对BMP信号的能力。在此筛选中鉴定出的基因将被进一步表征，以将它们置于腹外侧中胚层诱导途径中。

DESCRIPTION (provided by applicant): In vertebrates, numerous tissues and organs are derived in whole, or in part, from the mesoderm, including the skeletal muscles, heart, skeleton and kidney. A complex network of genes regulates the progressive specification of these various mesoderm derivatives. In Xenopus, the mesoderm forms from a band of cells around the equator of the early gastrula embryo. In models of mesoderm patterning in Xenopus, the Spemann organizer has figured prominently, and is thought to specify mesodermal fates through its secretion of inducing factors. We have recently shown that a Spemann organizer-independent pathway establishes a pattern along the animal-vegetal axis of the mesoderm by the mid-gastrula stage. Two distinct domains can be observed in this axis. The animal domain expresses the gene brachyury and appears to be fated to become notochord and somite, while the vegetal domain is fated to become ventrolateral mesoderm. The goal of this proposal is to further elucidate the mechanisms that act to establish patterning along the animal/vegetal axis. In particular, we will investigate the role of the nodal growth factors in the induction of ventral mesoderm. Our preliminary results indicate that Xenopus nodal-related-2 (Xnr2)-expressing cells are restricted to the presumptive ventral mesoderm in the gastrula embryo. Lineage tracing experiments proposed here are aimed at rigorously testing the fate of these cells. Conclusively establishing the fates of cells within the early presumptive mesoderm, and correlating them with restricted gene expression patterns, is an essential first step in elucidating the mechanism and timing of specification of fates. We have also hypothesized that nodals plays an essential role in the induction of ventral mesoderm, and we will examine the activity of nodals in the gastrula embryo by use of dominant negative constructs and inhibitory binding proteins. Finally, our preliminary results show that the combination of Xnr2 signaling and low FGF activity, such as exists in the vegetal marginal zone, makes ectoderm explants differentiate as blood, a ventral mesoderm derivative, in response to bone morphogenetic proteins (BMP). We will use subtraction and expression cloning approaches to identify genes downstream of Xnr2 signaling that are responsible for altering the competence of ectoderm explants to BMP signaling. Genes that are identified in this screen will be further characterized to place them in the pathway of ventrolateral mesoderm induction.